The delivery of therapeutic agents to specific tissues or sites within a body presents a variety of challenges, particularly where local delivery of a high dose of an insoluble therapeutic agent to a specific tissue is desired. While it has been recognized that modification of therapeutic agents by conjugation to soluble polymers may aid in their localized delivery, this technique has required the presence, or introduction, of functionalities into the therapeutic agent to accommodate a linkage to the polymer vehicle. Where it is necessary to introduce functionality into a therapeutic agent by chemical modification to accommodate a linkage, characteristics of the therapeutic agent such as potency, half-life, and metabolism may be altered. There is continuing need for polymer conjugates that can solublize and deliver therapeutic agents to specific tissues without requiring functionalization of the therapeutic agent to accommodate linkage to the polymer vehicle. Moreover, targeting of therapeutics to specific tissues via linkage to a polymeric delivery vehicle may result in diminished side effects.
There are insufficient means for conjugation of therapeutic agents via quinone and carbonyl functionalities to polymeric vehicles that can release the therapeutic agent unaltered under in vivo conditions. It may be advantageous to solubilize and deliver such carbonyl-containing and quinone-containing therapeutic agents to specific tissues by conjugation with suitable polymer vehicles. Therapeutic agents containing carbonyl or quinone functionalities that might advantageously be delivered by suitable polymeric vehicles include lobeline, acebutolol, methyprylon, haloperidol, molindone, naloxone, oxycodone, methadone, ketanserin, tolmetin, ketoprofen, nabumetone, canrenone, canrenonate, mebendazole, oxolinic acid, tetracycline, chlortetracycline, oxytetracycline, demeclocycline, doxycycline, minocycline, daunorubicin, doxorubicin, mitoxantrone, plicamycin, mitomycin, indan-1,3-dione, anisindione, testosterone (and related C-17 esters, e.g., propionate, enanthate, cypionate), dihydrotesterone, cyproterone acetate, estrone, progesterone, medroxyprogesterone acetate, hydroxyprogesterone caproate, norethindrone, norethynodrel, megestrol acetate, norgestrel, mifepristone, methandrostenolone, oxandrolone, testolactone, cyproterone acetate, prednisone, prednisolone, betamethasone, dexamethasone, other 3-, 17-, or 20-ketosteroids (e.g., dehydroepiandorsterone, androstenedione, cortisol, cortisone, aldosterone, etc.), and particularly β-lapachone compounds.
β-lapachone (3,4-dihydro-2,2-dimethyl-2H-naphtho[1,2-b]pyran-5,6-dione), a quinone, is derived from lapachol (a naphthoquinone) which can be isolated from the lapacho tree (Tabebuia avellanedae), a member of the catalpa family (Bignoniaceae). Lapachol and β-lapachone (with numbering) have the following chemical structures:

β-lapachone, as well as its intermediates, derivatives and analogs thereof, are described in Li, C. J. et al., (1993) J. Biol. Chem., 268(30): 22463-22468.
As a single agent, β-lapachone has demonstrated significant antineoplastic activity against human cancer cell lines at concentrations typically in the range of 1-10 μM (IC50). Cytotoxicity has been demonstrated in transformed cell lines derived from: patients with promyelocytic leukemia (Planchon et al., (1996) Cancer Res., 55: 3706-3711), prostate (Li, C. J., et al., (1995) Cancer Res., 55: 3712-3715), malignant glioma (Weller, M. et al., (1997) Int. J. Cancer, 73: 707-714), hepatoma (Lai, C. C., et al., (1998) Histol Histopathol, 13: 89-97), colon (Huang, L., et al., (1999) Mol Med, 5: 711-720), breast (Wuertzberger, S. M., et al., (1998) Cancer Res., 58: 1876), ovarian (Li, C. J. et al., (1999) Proc. Natl. Acad. Sci. USA, 96(23): 13369-13374), pancreatic (Li, Y., et al., (2000) Mol Med, 6: 1008-1015; Li, Y., (1999) Mol Med, 5: 232-239), and multiple myeloma cell lines, including drug-resistant lines (Li, Y., (2000) Mol Med, 6: 1008-1015). No cytotoxic effects were observed on normal fresh or proliferating human peripheral blood mononuclear cells (PBMC) (Li, Y., (2000) Mol Med, 6: 1008-1015).
β-lapachone appears to work by inducing unscheduled expression of checkpoint molecules, e.g., E2F, independent of DNA damage and cell cycle stages. Several studies have shown that β-lapachone activates checkpoints and induces cell death in cancer cells from a variety of tissues without affecting normal cells from these tissues (U.S. Patent Application Publication No. 2002/0169135). In normal cells with their intact regulatory mechanisms, such an imposed expression of a checkpoint molecule results in a transient expression pattern and causes little consequence. In contrast, cancer and pre-cancer cells have defective mechanisms, which result in unchecked and persistent expression of unscheduled checkpoint molecules, e.g., E2F, leading to selective cell death in cancer and pre-cancer cells.
β-lapachone has been shown to be a DNA repair inhibitor that sensitizes cells to DNA-damaging agents including radiation (Boothman, D. A. et al., Cancer Res, 47 (1987) 5361; Boorstein, R. J., et al., Biochem. Biophys. Commun., 117 (1983) 30). β-lapachone has also shown potent in vitro inhibition of human DNA Topoisomerases I (Li, C. J. et al., J. Biol. Chem., 268 (1993) 22463) and II (Frydman, B. et al., Cancer Res., 57 (1997) 620) with novel mechanisms of action. Unlike topoisomerase “poisons” (e.g., camptothecin, etoposide, doxorubicin) which stabilize the covalent topoisomerase-DNA complex and induce topoisomerase-mediated DNA cleavage, β-lapachone interacts directly with the enzyme to inhibit catalysis and block the formation of cleavable complex (Li, C. J. et al., J. Biol. Chem., 268 (1993) 22463), or β-lapachone interacts with the complex itself, causing religation of DNA breaks and dissociation of the enzyme from DNA (Krishnan, P. et al., Biochem Pharm, 60 (2000) 1367). β-lapachone and its derivatives have also been synthesized and tested as anti-viral and anti-parasitic agents (Goncalves, A. M., et al., Mol. Biochem. Parasitology, 1 (1980) 167-176; Schaffner-Sabba, K., et al., J. Med. Chem., 27 (1984) 990-994).
More specifically, β-lapachone appears to work by disrupting DNA replication, causing cell-cycle delays in G1 and/or S phase, inducing cell death in a wide variety of human carcinoma cell lines without DNA damage and independent of p53 status (Li, Y. Z. et al. (1999); Huang, L. et al.). Topoisomerase I is an enzyme that unwinds the DNA that makes up the chromosomes. The chromosomes must be unwound in order for the cell to use the genetic information to synthesize proteins; β-lapachone keeps the chromosomes wound tight, so that the cell cannot make proteins. As a result, the cell stops growing. Because cancer cells are constantly replicating and circumvent many mechanisms that restrict replication in normal cells, they are more vulnerable to topoisomerase inhibition than are normal cells.
Another possible intracellular target for β-lapachone in tumor cells is the enzyme NAP(P)H:quinone oxidoreductase (NQO1). Biochemical studies suggest that reduction of β-lapachone by NQO1 leads to a “futile cycling” between the quinone and hydroquinone forms with a concomitant loss of reduced NADH or NAD(P)H (Pink, J. J. et al., J. Biol. Chem., 275 (2000) 5416). The exhaustion of these reduced enzyme cofactors may be a critical factor for the activation of cell death pathways after β-lapachone treatment. Reinicke et al. reach a similar conclusion using mono(arylimino) derivatives of β-lapachone (Reinicke et al., Clin. Cancer. Res. 11(8) (2005) 3055-64).
As a result of these findings, β-lapachone is actively being developed for the treatment of cancer and tumors. In WO 00/61142, for example, there is disclosed a method and composition for the treatment of cancer, which comprises the administration of an effective amount of a first compound, a G1 or S phase drug, such as a β-lapachone, in combination with a G2/M drug, such as a taxane derivative. Additionally, U.S. Pat. No. 6,245,807 discloses the use of β-lapachone, amongst other β-lapachone derivatives, for use in the treatment of human prostate disease.
In addition to β-lapachone, a number of β-lapachone analogs having anti-proliferative properties have been disclosed in the art, such as those described in PCT International Application PCT/US93/07878 (WO 94/04145), and U.S. Pat. No. 6,245,807, in which a variety of substituents may be attached at positions 3- and 4- on the β-lapachone compound. PCT International Application PCT/US00/10169 (WO 00/61142), discloses β-lapachone, which may have a variety of substituents at the 3-position as well as in place of the methyl groups attached at the 2-position. U.S. Pat. Nos. 5,763,625, 5,824,700, and 5,969,163 disclose analogs with a variety of substituents at the 2-, 3- and 4-positions. Furthermore, a number of journals report β-lapachone analogs with substituents at one or more of the following positions: 2-, 3-, 8- and/or 9-positions. See, e.g., Sabba et al., (1984) J Med Chem 27:990-994 (substituents at the 2-, 8- and 9-positions); Portela and Stoppani, (1996) Biochem Pharm 51:275-283 (substituents at the 2- and 9-positions); Goncalves et al., (1998) Molecular and Biochemical Parasitology 1:167-176 (substituents at the 2- and 3-positions).
Moreover, structures having sulfur-containing hetero-rings in the “α” and “β” positions of lapachone have been reported (Kurokawa S, (1970) Bulletin of The Chemical Society of Japan 43:1454-1459; Tapia, R A et al., (2000) Heterocycles 53(3):585-598; Tapia, R A et al., (1997) Tetrahedron Letters 38(1):153-154; Chuang, C P et al., (1996) Heterocycles 40(10):2215-2221; Suginome H et al., (1993) Journal of the Chemical Society, Chemical Communications 9:807-809; Tonholo J et al., (1988) Journal of the Brazilian Chemical Society 9(2): 163-169; and Krapcho A P et al., (1990) Journal of Medicinal Chemistry 33(9):2651-2655). More particularly, hetero β-lapachone analogs are disclosed in PCT/US2003/037219, which published as WO 04/045557.
One obstacle to the development of pharmaceutical formulations comprising β-lapachone or β-lapachone analogs for pharmaceutical use is the low solubility of β-lapachone compounds in pharmaceutically acceptable solvents. β-lapachone compounds are generally highly insoluble in water and have only limited solubility in common solvent systems used for pharmaceutical administration, specifically for intravenous delivery of drugs. As a result, there is a need for improved formulations of β-lapachone compounds for pharmaceutical administration, which are both safe and readily bioavailable to the subject to which the formulation is administered. Importantly, there is an additional need to provide compositions that target tumor tissue with anti-cancer agents such as β-lapachone in such a manner that reduces the potential side effects due to the agent being released at unwanted sites. This invention describes systems for the delivery of therapeutic agents having carbonyl or quinone functionalities, including β-lapachone compounds, to tumors in a manner that diminishes unwanted side effects. As the polymeric compositions and methods described herein bind carbonyl-containing or quinone-containing therapeutic agents through a cleavable linkage to the carbonyl or quinone functionalities, they can advantageously deliver the therapeutic agents without the introduction of additional functionalities that may alter the structure, function, activity, or metabolism of the released therapeutic agents.